Newly observed binding mode in pancreatic ribonuclease

Aguilar, Carlos Fernando; Thomas, P.J.; Mills, Alberto; Moss, David S.; Palmer, Rex A.

doi:10.1016/0022-2836(92)90589-c

Cited by 38 publications

(31 citation statements)

References 7 publications

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“…This is in contrast to the results found for the complexes of RNase A with 2',5'-CpG and 3',5'-d(CpG) (Aguilar et al, 1991(Aguilar et al, , 1992Lisgarten et al, 1995), according to which the inhibitors are bound in a nonproductive way (retro-binding in the words of the authors) in which the downstream guanine is bound at the major binding site. As concluded by Zegers et a].…”

Section: Discussioncontrasting

confidence: 84%

Three‐dimensional structure of the complexes of ribonuclease A with 2′,5′‐CpA and 3′,5′‐d(CpA) in aqueous solution, as obtained by NMR and restrained molecular dynamics

et al. 1996

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The three‐dimensional structure of the complexes of ribonuclease A with cytidyl‐2′,5′‐adenosine (2′,5′‐CpA) and deoxycytidyl‐3′,5′‐deoxyadenosine [3′,5′‐d(CpA)] in aqueous solution has been determined by 1H NMR methods in combination with restrained molecular dynamics calculations. Twenty‐three intermolecular NOE cross‐correlations for the 3′,5′‐d(CpA) complex and 19 for the 2′,5′‐CpA, together with about 1,000 intramolecular NOEs assigned for each complex, were translated into distance constraints and used in the calculation. No significant changes in the global structure of the enzyme occur upon complex formation. The side chains of His 12, Thr 45, His 119, and the amide backbone group of Phe 120 are involved directly in the binding of the ligands at the active site. The conformation of the two bases is anti in the two complexes, but differs from the crystal structure in the conformation of the two sugar rings in 3′,5′‐d(CpA), shown to be in the S‐type region, as deduced from an analysis of couplings between the ribose protons. His 119 is found in the two complexes in only one conformation, corresponding to position A in the free protein. Side chains of Asn 67, Gln 69, Asn 71, and Glu 111 form transient hydrogen bonds with the adenine base, showing the existence of a pronounced flexibility of these enzyme side chains at the binding site of the downstream adenine. All other general features on the structures coincide clearly with those observed in the crystal state.

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Section: Discussioncontrasting

confidence: 84%

Three‐dimensional structure of the complexes of ribonuclease A with 2′,5′‐CpA and 3′,5′‐d(CpA) in aqueous solution, as obtained by NMR and restrained molecular dynamics

et al. 1996

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“…Indeed, in the ligand-free form, water molecules are located in the active site ( Figure 2B), whereas in lb-NCD-PLCC, dCpdG is positioned in the retrobinding mode similar to that observed for guanine containing dinucleotide complexed to RNase A [44][45][46] and BS-RNase. 47 The purine base is located in B1 1 and is hydrogen bonded to Thr45, the guanine ribose moiety is exposed to the solvent and does not interact with the protein, and the phosphate group is only partially visible.…”

Section: Ligands Bound At the Active Sitesmentioning

confidence: 84%

Toward an antitumor form of bovine pancreatic ribonuclease: The crystal structure of three noncovalent dimeric mutants

et al. 2009

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The cytotoxic action of bovine seminal ribonuclease (BS-RNase) depends on its noncovalent swapped dimeric form (NCD-BS), which presents a compact structure that allows the molecule to escape ribonuclease inhibitor (RI). A key role in the acquisition of this structure has been attributed to the concomitant presence of a proline in position 19 and a leucine in position 28. The introduction of Leu28, Cys31, and Cys32 and, in addition, of Pro19 in the sequence of bovine pancreatic ribonuclease (RNase A) has produced two dimeric variants LCC and PLCC, which do exhibit a cytotoxic activity, though at a much lower level than BS-RNase. The crystal structure analysis of the noncovalent swapped form (NCD) of LCC and PLCC, complexed with the substrate analogue 2 '-deoxycytidylyl(3 ',5 ')-2 '-deoxyguanosine, has revealed that, differently from NCD-BS, the dimers adopt an opened quaternary structure, with the two Leu residues fully exposed to the solvent, that does not hinder the binding of RI. Similar results have been obtained for a third mutant of the pancreatic enzyme, engineered with the hinge peptide sequence of the seminal enzyme (residues 16-22) and the two cysteines in position 31 and 32, but lacking the hydrophobic Leu residue in position 28. The comparison of these three structures with those previously reported for other ribonuclease swapped dimers strongly suggests that, in addition to Pro19 and Leu28, the presence of a glycine at the N-terminal end of the hinge peptide is also important to push the swapped form of RNase A dimer into the compact quaternary organization observed for NCD-BS.

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“…The structure of RNase A complexed with d(CpA) was primarily determined in order to gain more detailed information on the interactions at the downstream binding site and to compare the binding mode to that found in the crystal structures of RNase A soaked with 2',5'-CpG and d(CpG) (Aguilar et al, 1991(Aguilar et al, , 1992. In those last 2 high-resolution structures, the downstream guanine was bound in the major binding site, and the active site was occupied by an inorganic sulfate or phosphate anion.…”

Section: Discussionmentioning

confidence: 99%

“…Previously published structures of dinucleotide complexes of RNase A (UpcA, Richards & Wyckoff, 1973;2',5'-CpA, Wodak et al, 1977; 2'F-dUpA, Pavlovsky et al, 1978) were not determined at high resolution nor extensively refined. The recently published, highly refined structures of the complexes of RNase A with 2',5'-CpG and d(CpG) do not address this question because the inhibitors were found to be bound in a nonproductive way (Aguilar et al, 1991(Aguilar et al, , 1992.…”

Section: 'mentioning

confidence: 99%

The structures of rnase a complexed with 3′‐CMP and d(CpA): Active site conformation and conserved water molecules

et al. 1994

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The interactions of RNase A with cytidine 3'-monophosphate (3"CMP) and deoxycytidyl-3',5'-deoxyadenosine (d(CpA)) were analyzed by X-ray crystallography. The 3'-CMP complex and the native structure were determined from trigonal crystals, and the d(CpA) complex from monoclinic crystals. The differences between the overall structures are concentrated in loop regions and are relatively small. The protein-inhibitor contacts are interpreted in terms of the catalytic mechanism. The general base His 12 interacts with the 2' oxygen, as does the electrostatic catalyst Lys 41. The general acid His 119 has 2 conformations (A and B) in the native structure and is found in, respectively, the A and the B conformation in the d(CpA) and the 3'-CMP complex. From the present structures and from a comparison with RNase T1, we propose that His 119 is active in the A conformation. The structure of the d(CpA) complex permits a detailed analysis of the downstream binding site, which includes His 119 and Asn 71. The comparison of the present RNase A structures with an inhibitor complex of RNase T1 shows that there are important similarities in the active sites of these 2 enzymes, despite the absence of any sequence homology. The water molecules were analyzed in order to identify conserved water sites. Seventeen water sites were found to be conserved in RNase A structures from 5 different space groups. It is proposed that 7 of those water molecules play a role in the binding of the N-terminal helix to the rest of the protein and in the stabilization of the active site.

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Newly observed binding mode in pancreatic ribonuclease

Cited by 38 publications

References 7 publications

Three‐dimensional structure of the complexes of ribonuclease A with 2′,5′‐CpA and 3′,5′‐d(CpA) in aqueous solution, as obtained by NMR and restrained molecular dynamics

Three‐dimensional structure of the complexes of ribonuclease A with 2′,5′‐CpA and 3′,5′‐d(CpA) in aqueous solution, as obtained by NMR and restrained molecular dynamics

Toward an antitumor form of bovine pancreatic ribonuclease: The crystal structure of three noncovalent dimeric mutants

The structures of rnase a complexed with 3′‐CMP and d(CpA): Active site conformation and conserved water molecules

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